| 质粒名称: | cytoplasmic EKAR (Cerulean-Venus) |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 18679 |
| 存储实验室: | Karel Svoboda |
|
载体骨架基本信息 |
|
| 载体名称: | pRK5 |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 4500 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达 |
| 筛选标记: | -- |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | ERK Activity Reporter,EKAR |
| 物种来源: | -- |
| 插入大小: | 550 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: |
monomeric Cerulean（插入件上的N端）
monomeric Venus（插入件上的C端子）
|
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | CMV Forward |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | cytoplasmic EKAR (Cerulean-Venus) was a gift from Karel Svoboda (Addgene plasmid # 18679 ; http://n2t.net/addgene:18679 ; RRID:Addgene_18679) |
| 参考文献部分: | A genetically encoded fluorescent sensor of ERK activity. Harvey CD, Ehrhardt AG, Cellurale C, Zhong H, Yasuda R, Davis RJ, Svoboda K.. PNAS 2008 Dec 9;105(49):19264-9 10.1073/pnas.0804598105 PubMed 19033456 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
质粒图.png)
载体简介
载体序列
LOCUS cytoplasmic EKAR (Cerulean-Venus) 6758 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS cytoplasmic EKAR (Cerulean-Venus)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6758)
AUTHORS Harvey CD, Ehrhardt AG, Cellurale C, Zhong H, Yasuda R, Davis RJ,
Svoboda K.
TITLE A genetically encoded fluorescent sensor of ERK activity
JOURNAL PNAS 2008 Dec 9;105(49):19264-9
PUBMED 19033456
REFERENCE 2 (bases 1 to 6758)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6758
/organism="synthetic DNA construct"
/mol_type="other DNA"
enhancer 14..393
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 394..597
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 548..568
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
promoter 827..845
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 827..844
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind 858..877
/label=pMT2-F
/note="Synthetic intron, forward primer"
regulatory 933..942
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 939..1655
/codon_start=1
/product="enhanced monomeric variant of CFP (Rizzo et al.,
2004)"
/label=mCerulean
/note=" mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS 2220..2936
/codon_start=1
/product="Venus YFP with monomerizing A206K mutation (Nagai
et al., 2002; Kremers et al., 2006)"
/label=mVenus
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
regulatory 3007..3016
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
polyA_signal 3020..3154
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(3057..3076)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 3111..3130
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
promoter 3223..3580
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
primer_bind complement(3246..3266)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 3431..3566
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 3493..3512
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(3600..3617)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(3600..3616)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3609..3631)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 3829..4284
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(3916..3935)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4126..4147
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4301..4320)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 4455..4477
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(4530..4548)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 4616..4720
/gene="bla"
/label=AmpR promoter
CDS 4721..5581
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(4939..4958)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5752..6340
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6241..6260
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 6494..6511
/label=L4440
/note="L4440 vector, forward primer"
promoter 6664..6694
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6702..6718
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 6707..6729
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 6726..6742
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 6726..6742
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
ORIGIN
1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
601 ttagtgaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagac
661 accgggaccg atccagcctc cgcggccggg aacggtgcat tggaacgcgg attccccgtg
721 ccaagagtga cgtaagtacc gcctatagag tctataggcc cacccccttg gcttcgttag
781 aacgcggcta caattaatac ataaccttat gtatcataca catacgattt aggtgacact
841 atagaataac atccactttg cctttctctc cacaggtgtc cactcccagg tccaactgca
901 cctcggttct atcgattgaa ttcgtaccgg tcgccaccat ggtgagcaag ggcgaggagc
961 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
1021 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
1081 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctggg
1141 gcgtgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
1201 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
1261 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
1321 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aacgccatca
1381 gcgacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc aacttcaaga
1441 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
1501 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccaagc
1561 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
1621 ccgggatcac tctcggcatg gacgagctgt acaagcggat ccatatggcg gacgaggaga
1681 agctgccgcc cggctgggag aagcgcatga gccgcagctc aggccgagtg tactacttca
1741 accacatcac taacgccagc cagtgggagc ggcccagcgg caacagcagc agtggtggca
1801 aaaacgggca gggggagcct gccaggaagc ttcgaattct gcagtcgact agggagcaag
1861 ggcttgtgtc tagaggcgga ggtggtggag gaggtggagg tggtggaggt ggaggtggag
1921 gtggtggcgg tggtggagga ggaggagcta gaggcggagg tggtggagga ggtggaggtg
1981 gtggaggtgg aggtggaggt ggtggcggtg gtggaggagg aggagctagc ggcggaggtg
2041 gtggaggagg tggaggtggt ggaggtggag gtggaggtgg tggcggtggt ggaggaggag
2101 gaatggctga ccaactgact gaagagtggt cgacggtacc gcggaccggt cctgacgtac
2161 cacgtacacc agtaggaaag gcaaagctgt cattccaatt cccgcgggcc cgggatatca
2221 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
2281 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
2341 gcaagctgac cctgaagctg atctgcacca ccggcaagct gcccgtgccc tggcccaccc
2401 tcgtgaccac cctgggctac ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc
2461 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
2521 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
2581 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
2641 agctggagta caactacaac agccacaacg tctatatcac cgccgacaag cagaagaacg
2701 gcatcaaggc caacttcaag atccgccaca acatcgagga cggcggcgtg cagctcgccg
2761 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
2821 acctgagcta ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc
2881 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaaggatg
2941 ctcttcaaaa aaaacttgaa gaacttgaac ttgacgaatg atctagagtc gacctgcaga
3001 agcttggccg ccatggccca acttgtttat tgcagcttat aatggttaca aataaagcaa
3061 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
3121 caaactcatc aatgtatctt atcatgtctg gatcgggaat taattcggcg cagcaccatg
3181 gcctgaaata acctctgaaa gaggaacttg gttaggtacc ttctgaggcg gaaagaacca
3241 gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
3301 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
3361 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
3421 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
3481 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
3541 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctgttaa cagcttggca
3601 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
3661 cttgcagcac atcccccctt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
3721 ccttcccaac agttgcgtag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt
3781 acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta
3841 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
3901 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
3961 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
4021 acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat
4081 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
4141 aaactggaac aacactcaac cctatctcgg gctattcttt tgatttataa gggattttgc
4201 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
4261 acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg
4321 catagttaag ccaactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
4381 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
4441 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
4501 cgcgaggcag tattcttgaa gacgaaaggg cctcgtgata cgcctatttt tataggttaa
4561 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
4621 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
4681 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
4741 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
4801 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
4861 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
4921 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgatgacg ccgggcaaga
4981 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
5041 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
5101 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
5161 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
5221 gaatgaagcc ataccaaacg acgagcgtga caccacgatg ccagcagcaa tggcaacaac
5281 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
5341 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
5401 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
5461 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
5521 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
5581 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
5641 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
5701 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
5761 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
5821 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
5881 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
5941 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
6001 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
6061 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
6121 actgagatac ctacagcgtg agcattgaga aagcgccacg cttcccgaag ggagaaaggc
6181 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
6241 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
6301 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
6361 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
6421 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
6481 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc
6541 gcctctcccc gcgcgttggc cgattcatta atccagctgg cacgacaggt ttcccgactg
6601 gaaagcgggc agtgagcgca acgcaattaa tgtgagttac ctcactcatt aggcacccca
6661 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
6721 tcacacagga aacagctatg accatgatta cgaattaa
//
